The soluble storage carbohydrate found in plants, including sucrose, glucans, starch and fructans, are an important source of feed for animals, particularly grazing ruminants. These carbohydrates are stored non-structurally which makes them readily available for digestion by animals and therefore an important source of digestible energy.
During periods of high sink activity and low source activity, such as during a drought, the level of stored carbohydrates falls as the non-structural storage carbohydrates are mobilised for use in seed filling. The result of this mobilisation, particularly in relation to pasture grasses, is a significant loss of feed value to grazing ruminants due to the reduction in the levels of the stored carbohydrates. This reduction is caused by the enzymatic degradation of the stored carbohydrates. This enzymatic degradation is assisted by the fact that the stored carbohydrates generally have a low degree of polymerization. For example, as noted by Radojevic et al 1994, during the period from late spring to early autumn in southern Australia, the declining feed quality of the grasses causes a corresponding reduction in the lactation by dairy herds and necessitates the use of supplementary feeds. This decline in digestibility is associated with a decline in the level of soluble carbohydrates.
Perennial rye grass lines which accumulate high concentrations of soluble carbohydrates from late spring to early autumn do not suffer as large a decline in digestibility (Radojevic et al 1994). The result of this increased digestibility is a corresponding increase in milk production by dairy herds.
In addition to this, there are many pasture plants, such as white clever which do not possess any significant levels of stored carbohydrate.
There has, therefore, been a desire to develop methods for preventing the degradation of the stored carbohydrates during plant senescence and to increase the level of stored carbohydrates in pasture plants with low levels.
Glucosyltransferases of Streptococcus salivarius
It is known that many strains of Streptococcus salivarius and Streptococcus mutans, produce extracellular .alpha.-D-glucosyltransferase (Gtfs), an enzyme which catalyses the formation of glucan from sucrose. These Gtfs are also found in many other species of oral streptococci.
The Gtfs utilise the high free energy of the glycosidic bond of sucrose to synthesise glucans (Jacques N A, Giffard P M, 1991). Gtfs produce either soluble or insoluble products by transferring a glucose residue from sucrose to a growing glucan chain.
Gtfs which produce an insoluble product are generally considered to be primer-dependent (Walker G J, Jacques N A, 1987). These primer-dependent Gtfs require a dextran (.alpha.-(1.fwdarw.6)-linked glucan) as a receptor for polymerisation to proceed at an appreciable rate. In contrast, Gtfs that produce soluble products may be either primer-dependent or primer-independent. The genetic sequences for 10 gtf genes from a number of Streptococcus species have been ascertained (Gilmore K S, Russell R R B, Ferretti J J). All the Gtfs coded by these genes possess highly conserved putative signal sequences that lead to the secretion of these enzymes. The remainder of each protein is arbitrarily divided into two domains--the N-terminal two-thirds "catalytic domain" and the C-terminal one-third "glucan-binding domain".
S. salivarius ATCC 25975 has been shown to possess at least four different gtf genes (Giffard et al (1991); Giffard et al (1993)). Each of these genes codes for a highly hydrophilic monomeric glucosyltransferase that possesses unique enzymic properties. These Gtfs synthesize structurally different glucans from sucrose. For example, the genes coding for GtfJ and GtfL produce enzymes which synthesize insoluble glucans. GtfJ is a primer-dependent enzyme producing essentially a linear a(1.fwdarw.3)-glucan while GtfL is a primer-independent enzyme that synthesizes a glucan containing 50% .alpha.-(1.fwdarw.3)--and 50% .alpha.-(1.fwdarw.6)-linked glucosyl residues. In contrast, the gtfK and gtfM genes code for enzymes which produce a soluble glucan which possess .alpha.-(1.fwdarw.6)-linked glucosyl residues. GtfK is primer stimulated while GtfM is primer independent.